Effect of growth development temperature on Cuticular Hydrocarbon (CHC) profiles of Spiniphora genitalis 3rd instar larvae

In forensic entomology, precise estimation of the postmortem interval (PMI) is important in death investigation, particularly when decomposition exceeds 72 hours. Analysis involving cuticular hydrocarbons (CHCs) is one of promising biochemical approach which have been shown to vary according to insect developmental stage and environmental conditions. This study investigates the effect of growth development temperature on the CHC profiles of Spiniphora genitalis 3rd instar larvae using Gas Chromatography-Mass Spectroscopy (GC-MS) and Principal Component Analysis (PCA). Larvae were reared at three different temperatures (21oC, 23oC, 25oC) and CHCs were extracted using hexane and analyzed via GC-MS. The hydrocarbon compound was identified using library search (NIST08), and electron ionization in scan mode was applied for mass spectroscopic detection. The resulting hydrocarbon profiles were subjected to PCA to assess temperature induced variation. The GC-MS studies demonstrated that 30 hydrocarbon were present in cuticle of 3rd instar larvae Spiniphora genitalis across different development temperature where 5 of it which is hexadecane, 2,4-Di-tert-butylphenol, Heptadecane, Octadecane and Heneicosane were present in all temperature. At least one hydrocarbon specifically present on each development temperature. This showed that every development temperature can be distinguish based on these unique hydrocarbon that only present on each specific temperature. There is also increase numbers of long hydrocarbon chain specifically straight chain alkane with temperature increase as a response to environmental stressor (temperature) due to responsible of it to minimize water loss thus prevent desiccation. PCA successfully cluster CHC profiles of 3rd instar larvae Spiniphora genitalis develop at different temperature. This findings indicate that CHCs could be used as reliable tools for development temperature identification of 3rd instar larvae. For future studies it is recommended to increase the replicates of the sample because PCA is more reliable when involving a large group of data set and must incorporate additional developmental stages and both sexes of Spiniphora genitalis to determine whether similar temperature-dependent CHC patterns are observed throughout the life cycle and across genders.